This application claims the benefit of Japanese Applications No. 2000-105968 and No. 2001-109394, which are hereby incorporated by reference.
1. Field of the Invention
The present invention relates generally to a minute particle optical manipulation method and a minute particle optical manipulation apparatus, and more particularly to a minute particle optical manipulation method and a minute particle optical manipulation apparatus for three-dimensionally trapping and moving a minute particle by irradiating the minute particle with beams.
2. Related Background Art
A technology of optically manipulating a minute particle is generally known as optical tweezers and optical trapping. This technology involves the use of mainly a laser and is therefore called laser trapping and laser tweezers.
This technology is that laser beam emitted from a radiation source is converged in a conical shape by a converging optical system and falls upon the vicinity of the minute particle existing in the medium, and the minute particle is trapped or held and moved by making use of a radiation pressure occurred about the minute particle. This technology is utilized in diversity as a method of trapping and manipulating a cell of a living body and a microbe in a non-contact and non-destructive manner.
An explanation of how the minute particle is manipulated by the optical tweezers described above will be made referring to FIGS. 11 and 12. Herein, FIG. 11 is a view schematically showing a configuration of the prior art optical tweezers. FIG. 12 is a partially enlarged view of FIG. 11 and explanatorily shows how a minute particle S is manipulated by the optical tweezers.
As illustrated in FIG. 11, a parallel beam L11 for the optical tweezers, which is emitted from a light source LS1 for the optical tweezers, is reflected in a wavelength-selective manner by a dichroic mirror DM and enters a normal converging optical system O3 of which a spherical aberration is substantially zero. Then, the vicinity of the minute particle S in a medium B held by a holder H such as a Petri dish and a slide glass, is irradiated with a cone-shaped converged beam L13 with no spherical aberration, which has passed through the converging optical system O3.
Note that mainly a laser is herein used as the optical tweezers oriented light source LS1, and an objective lens for a transmission type optical microscope (that is hereinafter simply called a xe2x80x9cmicroscope objective lensxe2x80x9d) is often used in terms of utilization as the converging optical system O3.
Thus, as shown in FIG. 12, if the minute particle S exists in the vicinity of a converging point P at which the beam is converged in the conical shape by the converging optical system O3, this cone-shaped converged beam L13 is reflected by the surface of the minute particle S and refracted inside the minute particle S, thus deflecting its traveling direction. As a result, a beam momentum changes. At this time, a radiation pressure corresponding to a change in the momentum of the converged beam L13 occurs about the minute particle S, and there acts a force F as indicated by a bold solid line in FIG. 12.
Now, supposing that the minute particle S has a refractive index higher than that of the medium B surrounding the particle S and is classified as a non-absorptive spherical minute particle, it is known from an analysis of the change in the momentum of the converged beam that the radiation pressure acts toward a higher light intensity, and the force F acts to get the minute particle F attracted to the converging point P. Accordingly, it is feasible to trap and manipulate the minute particle S by making use of this force F.
Further, when thus trapping and manipulating the minute particle S in the medium B, it is necessary to observe how the particle is trapped and manipulated, and therefore an observation optical system is provided.
Namely, illumination beam L2 emitted from an observation light source LS2 provided under the holder H travels through an illumination optical system C1 and illuminates over the vicinity of the minute particle S in the medium B. Thereafter, the illumination beam L2 passes through the converging optical system O3, then penetrates a dichroic mirror DM and is projected on an image surface IMG to form an image thereon.
Then, an enlarged image of the minute particle S that is formed on this image surface IMG is viewed by a naked eye E through an imaging device D such as a CCD camera etc. as well as through an eyepiece EP, thereby making it possible to observe how the minute particle S in the medium B is trapped and manipulated.
In the conventional optical tweezers, however, it is known that a force for trapping the minute particle S in an axial direction, i.e., an optical-axis direction (which will hereinafter referred to as a xe2x80x9ctrapping forcexe2x80x9d) along a traveling direction of the optical tweezers oriented converged beam L13, is by far smaller than a trapping force acting in a direction perpendicular to the optical-axis direction.
It is generally known that beam containing a component having a larger angle to the optical axis, i.e., a high NA (Numerical Aperture) component, is useful for obtaining a strong trapping force in the optical tweezers. In fact, however, it is difficult in terms of optics to actualize the converged beam having a numerical aperture of 1.5 or larger (NA=1.5 or above). Further, there is also a method of strengthening the light intensity with which the minute particle is irradiated, however, if a large output light source is used, there might be a possibility in which a minute living sample is damaged or destructed.
Therefore, what is desired is a method of enhancing the trapping force in the optical-axis direction without strengthening the intensity of the beam irradiation upon the minute particle, and as a matter of fact some proposals have been made so far.
xe2x80x9cLaser Trapping Method and Apparatusxe2x80x9d (Japanese Patent No. 2947971) and xe2x80x9cLaser Trapping Apparatus and Prism Used thereofxe2x80x9d (Japanese Patent Application Laid-Open No. 8-262328), may be given by examples thereof.
The laser trapping related to each of those proposals utilizes such a principle that the high NA component, having the large angle to the optical axis, of the converged beam makes a great contribution to the trapping force, while the component having a small angle does not contribute to the trapping force so much. The laser trapping is based on such a structure that a prism taking a special shape is inserted into the light path, parallel light beam from the light source is thereby converted into a converged beam taking a conical cylindrical shape that is composed of only a large angle component without any loss, and the sample is irradiated with the converged beam.
The laser trapping related to each of those proposals, however, involves inserting the specially-shaped prism into the light path in order to convert the beam into the conical cylindrical shape. Further, it is required that the beam substantially symmetric about the optical axis be obtained for stably trapping the sample, and hence there is a demand for a highly precise adjustment of a position of the prism.
As a result, each of the laser trapping apparatuses related to the proposals given above involves the use of an expensive prism element and therefore costs high. Another problem is that this laser trapping apparatus needs a mechanism for accurately holding the prism, which leads a scale-up of the apparatus.
Moreover, the trapping force in the optical-axis direction is enhanced because of using the conical cylindrical converged beam, however, a range where the trapping force acts in the optical-axis direction shrinks, resulting in a problem that only the sample in close proximity to the converging point can be trapped.
Accordingly, it is a target to actualize the optical tweezers capable of enhancing the trapping force in the optical-axis direction and expanding the range where the trapping force acts in the optical-axis direction without requiring an optical element such as a special prism etc.
Further, generally the optical tweezers have a comparatively weak trapping force in the optical-axis direction, and besides the range where the trapping force acts in the optical-axis direction is limited. Hence, in the case of trapping and manipulating the minute particle existing in a deep position in the medium, there exists a necessity of making an adjusting for getting tips of the optical tweezers, i.e., the converging point of the beam close to the vicinity of the minute particle, namely making a focusing adjustment.
In fact, however, there arises a problem in which even when making the focusing adjustment, the maximum trapping force obtained by the optical tweezers decreases as the position of the minute particle in the medium gets deeper, resulting in a difficulty of trapping the minute particle.
This is because a distance at which the beam travels through the medium surrounding the minute particle becomes longer as the position of the minute particle in the medium gets deeper, with the result that a minus spherical aberration occurs in the converged beam.
For instance, if the minute particle in the medium composed of a liquid such as water is trapped through a cover glass, an objective lens for observing a living body, which is often used as a converging optical system for the conventional optical tweezers, is adjusted so that the spherical aberration is zero at the under surface of the cover glass. If the beam is converged at a position deep within the medium under the cover glass, the minus spherical aberration occurs when passing through the medium. Therefore, the maximum trapping force obtained by the optical tweezers becomes weaker as the position of the minute particle in the medium gets deeper, and it is difficult to trap the minute particle. Further, the same situation also occurs in the case of trapping a molecule existing not in proximity to the surface of a thick living sample but in a position deep inside.
Accordingly, it has been a target to actualize the optical tweezers capable of obtaining the trapping force enough to trap the particle even when the minute particle exists deep within the medium.
It is a primary object of the present invention, which was devised to obviate the problems inherent in the prior art, to provide a minute particle optical manipulation method and a minute particle optical manipulation apparatus that are capable of simply strengthening a trapping force in an optical-axis direction and expanding a range where the trapping for acts in the optical-axis direction without requiring an optical element such as a special prism etc., and obtaining the trapping force enough to trap the particle even when the minute particle exists deep within a medium while keeping the trapping force when the minute particle is in a shallow position within the medium.
To accomplish the above object, the present inventor discovered that after calculating the trapping force in the optical-axis direction in each case of changing in many ways a condition of beam with which the minute particle in the medium is irradiated, the trapping force in the optical-axis direction is strengthened if a plus spherical aberration is intentionally given to a cone-shaped converged beam with which the minute particle in the medium is irradiated.
Then, as a result of having made examinations over and over in concentration on the basis of the above knowledge, it was confirmed that when irradiating the minute particle in the medium with the cone-shaped converged beam having the plus spherical aberration, the range where the trapping force acts in the optical-axis direction is expanded as well as strengthening the trapping force in the optical-axis direction, and a sufficiently strong trapping force is obtained even when the minute particle exists deep inside the medium.
Accordingly, the above object is accomplished by a minute particle optical manipulation method and a minute particle optical manipulation apparatus according to the present invention.
According to a first aspect of the present invention, a minute particle optical manipulation method comprises a step of irradiating a minute particle in a medium with a cone-shaped converged beam having a plus spherical aberration, and a step of trapping and manipulating the minute particle.
In the minute particle optical manipulation method according to the first aspect, the trapping force in the optical-axis direction is more strengthened and the range where the trapping force acts in the optical-axis direction is more expanded by irradiating the minute particle in the medium with the cone-shaped converged beam having the plus spherical aberration and trapping and manipulating the minute particle without making a high-level adjustment such as inserting a special prism than in a case of converging a cone-shaped converged beam having no aberration at one point.
Further, if the minute particle exists deep inside the medium under the cover glass, a minus spherical aberration occurs when the converged beam travels through the medium in the prior art. The minus spherical aberration occurred depending on a depth in the medium can be intentionally, however, canceled and converted into a plus spherical aberration by use of the cone-shaped converged beam having the plus spherical aberration. It is therefore feasible to obtain the sufficiently strong trapping force while keeping the trapping force when the minute particle exists shallow in the medium.
Note that a method of giving the plus spherical aberration to the cone-shaped converged beam with which the minute particle in the medium is irradiated involves the use of a converging optical system designed and manufactured so that the optical system itself has the plus spherical aberration. Other than this method, there are a variety of methods capable of simply generating the plus spherical aberration even when using the converging optical system having almost no occurrence of the spherical aberration as exemplified by an existing objective lens of a microscope.
For example, in the converging optical system that causes almost no spherical aberration, there are methods such as putting a transparent thin plane-parallel plate in a position where the beam on the light path diverge or converge and diverging or converging the beam, disposing a diffraction optical element for generating the spherical aberration on the light path, a method of changing an arranging interval (air spacing) by moving in the optical-axis direction some lenses of a lens unit constituting the converging optical system, replacing, when using a cover glass, this cover glass with one exhibiting a high refractive index, and exchanging, when using an oil-immersed objective lens, this oil with one having a high refractive index.
According to a second aspect of the present invention, a minute particle optical manipulation method according to the first aspect may further comprise a step of arbitrarily changing the plus spherical aberration of the cone-shaped converged beam in accordance with a condition of the minute particle in the medium.
In the minute particle optical manipulation method according to the second aspect, the plus spherical aberration of the cone-shaped converged beam with which the minute particle is irradiated is arbitrarily changed, whereby the optimum plus spherical aberration can be selected if the conditions of the target minute particle itself, e.g., a size and a material of the minute particle are different, and if the conditions under which the minute particle exists, e.g., a material of the medium and a depth in which the minute particle exists in the medium are different. Hence, the minute particle optical manipulation method according the first aspect yields effects wherein the trapping force in the optical-axis direction is strengthened, the range in which the trapping force acts in the optical-axis direction is expanded, and the sufficiently strong trapping force is obtained in the deep position in the medium while keeping the trapping force when the minute particle exists in a shallow position in the medium.
Note that the method of arbitrarily changing the plus spherical aberration of the cone-shaped converged beam with which the minute particle in the medium is irradiated may be, if exemplified corresponding to the method of giving the plus spherical aberration in the minute particle optical manipulation method according to the first aspect, for instance, a method of preparing plural types of transparent thin plane-parallel plates for diverging or converging the beam and diffraction optical elements for generating the spherical aberration, selecting those exhibiting desired characteristics and inserting or removing them in predetermined positions on the light path of the converging optical system with almost no occurrence of the spherical aberration, a method of changing the arranging interval (air spacing) by further moving in the optical-axis direction some lenses of the lens unit constituting the converging optical system, replacing, when using the cover glass, this cover glass with other cover glass exhibiting a different refractive index, and exchanging, when using the oil-immersed objective lens, this oil with other oil having a different refractive index.
In a minute particle optical manipulation method according to the first or second aspect, it is preferable that there be established a relationship such as:
n1 greater than n2 
where n1 is a refractive index of the minute particle, and n2 is a refractive index of the medium, and a spherical aberration SA with respect to a maximum NA component of the cone-shaped converged beam has the following relationship:
0.2 Rxe2x89xa6SAxe2x89xa61.5 R 
where R is a radius of the minute particle.
Then, more essentially, it is desirable in order to obtain the trapping force most effectively especially when the minute particle exists in a comparatively shallow position in the medium that the spherical aberration SA with respect to the maximum NA component of the cone-shaped converged beam has the following relationship:
0.2 Rxe2x89xa6SAxe2x89xa61.0 R 
Still further, it is more desirable in order to obtain the trapping force most effectively particularly when the minute particle exists in a comparatively deep position in the medium that the spherical aberration SA with respect to the maximum NA component of the cone-shaped converged beam has the following relationship:
0.75 Rxe2x89xa6SAxe2x89xa61.5 R 
According to a third aspect of the present invention, a minute particle optical manipulation apparatus comprises a converging optical system for generating a cone-shaped converged beam having a plus spherical aberration, wherein a minute particle in a medium is irradiated with the cone-shaped converged beam having the plus spherical aberration that emerges from the converging optical system, and is trapped and manipulated.
Thus, the minute particle optical manipulation apparatus according to the third aspect has the converging optical system for generating the cone-shaped converged beam having the plus spherical aberration. It is therefore possible to easily carry out the minute particle optical manipulation method according to the first aspect that includes the steps of irradiating the minute particle in the medium with the cone-shaped converged beam having the plus spherical aberration and trapping and manipulating the minute particle. Hence, there are exhibited the effects of the minute particle optical manipulation method according to the first aspect such as strengthening the trapping force in the optical-axis direction, expanding the range in which the trapping force acts in the optical-axis direction, and obtaining the sufficiently strong trapping force in the deep position in the medium while keeping the trapping force when the minute particle exists in a shallow position in the medium.
Note that the converging optical system for generating the cone-shaped converged beam having the plus spherical aberration may be herein a converging optical system designed and manufactured to have the plus spherical aberration from the beginning. Other than this optical system, however, there are a variety of converging optical systems each capable of easily generating the plus spherical aberration even if using the converging optical system with almost no occurrence of the spherical aberration as in the case of an existing microscope objective lens.
For instance, some of the converging optical systems with almost no occurrence of the spherical aberration have such a geometry that the transparent thin plane-parallel plate is disposed in the position for diverging or converging the beam on the light path, that the diffraction optical element for generating the spherical aberration is disposed on the light path, and that some lenses of the lens unit constituting the converging optical system are moved in the optical-axis direction to change the arranging interval (air spacing).
According to a fourth aspect of the present invention, a minute particle optical manipulation apparatus according to the third aspect may further comprise a spherical aberration changing device for arbitrarily changing the plus spherical aberration of the cone-shaped converged beam which is generated by the converging optical system in accordance with a condition of the minute particle in the medium.
Thus, the minute particle optical manipulation apparatus according to the fourth aspect includes the spherical aberration changing device for arbitrarily changing the plus spherical aberration of the cone-shaped converged beam generated by the converging optical system. It is therefore feasible to easily carry out the minute particle optical manipulation method according to the second aspect that includes the step of arbitrarily changing the plus spherical aberration of the cone-shaped converged beam in accordance with the condition of the minute particle in the medium. Hence, there exhibited the effects of the minute particle optical manipulation method according to the second aspect such as strengthening the trapping force in the optical-axis direction, expanding the range in which the trapping force acts in the optical-axis direction, and obtaining the sufficiently strong trapping force in the deep position in the medium while keeping the trapping force when the minute particle exists in a shallow position in the medium, corresponding to changes in the variety of conditions of the minute particle in the medium.
Note that as the spherical aberration changing device for arbitrarily changing the plus spherical aberration of the cone-shaped converged beam with which the minute particle in the medium is irradiated, if corresponding to the element for giving the plus spherical aberration as exemplified in the minute particle optical manipulation apparatus according to the third aspect, it may be considered to provide an inserting/removing mechanism wherein plural types of, e.g., transparent thin plane-parallel plates for diverging or converging the beam and diffraction optical elements for generating the spherical aberration are prepared, the plane-parallel plate and the diffraction optical element exhibiting desired characteristics are selected from those plates and elements and inserted in or removed from predetermined positions on the light path of the converging optical system with almost no occurrence of the spherical aberration, and a lens moving mechanism for moving some lenses of the lens unit constituting the converging optical system and further changing the arranging interval (air spacing) thereof.
According to a fifth aspect of the present invention, a minute particle optical manipulation apparatus according to the third or fourth aspect may further comprise an observation optical system, including a part of the whole of the converging optical system, for observing the minute particle, wherein the observation optical system is provided with a correcting mechanism for correcting the plus spherical aberration of the converging optical system or an in-focus position of the observation optical system.
Thus, in the minute particle optical manipulation apparatus according to the fifth aspect, the observation optical system containing a part of the whole of the converging optical system is provided with the correction mechanism for correcting the plus spherical aberration of the converging optical system or the in-focus position of the observation optical system. Therefore, the observation optical system shares a part or the whole of the converging optical system for generating the cone-shaped converged beam having the plus spherical aberration. Even if the spherical aberration and a defocus occur in the observation optical system due to the above configuration, the correction mechanism is capable of correcting the spherical aberration and the defocus, and it is therefore possible to prevent an occurrence of such a situation that an observed image of the minute particle is viewed in blur when observing the minute particle through the observation optical system with the result that only a low contrast is obtained.
According to a sixth aspect of the present invention, in a minute particle optical manipulation apparatus according to the third or fourth aspect, the observation optical system for observing the minute particle is provided independently of the converging optical system.
Thus, in the minute particle optical manipulation apparatus according to the sixth aspect of the present invention, the observation optical system is provided independently of the converging optical system, and hence it is feasible to avoid the spherical aberration and the defocus from occurring in the observation optical system because of sharing a part or the whole of the converging optical system. It is therefore possible to prevent an occurrence of such a situation that the observed image of the minute particle is viewed in blur when observing the minute particle through the observation optical system with the result that only the low contrast is obtained.
Next, in a minute particle optical manipulation apparatus according to a seventh aspect of the present invention, an axial chromatic aberration xcex94 of observation light on the basis of trapping light in the converging optical system is set to have a predetermined negative value. In this case, the converging position of a converged beam which is generated when parallel beams of the trapping light enter the converging optical system is farther from the converging optical system than the in-focus position with respect to the converging optical system which serves as an objective lens in the observation system only by a predetermined distance (that is, the axial chromatic aberration xcex94) along the optical axis of the converging optical system.
Consequently, in the minute particle optical manipulation apparatus according to the seventh aspect of the present invention, the converging position of the converged beam is moved toward the converging optical system along the optical axis in order to observe an excellent image of a minute particle with high contrast by making the position of the minute particle which is trapped by the action of the converged beam (and the converging position of the converged beam, in its turn) substantially coincident with the in-focus position. In this case, upon movement of the converging position of the converged beam toward the converging optical system, a plus spherical aberration is given to the converged beam. As a result, as will be described later, it is possible to maintain the trapping force (the force for trapping a minute particle) stably and strongly by the action of the converged beam with the plus spherical aberration given thereto. It is also possible to observe an excellent image of the minute particle with high contrast since the converging position of the converged beam is substantially coincident with the in-focus position.
Description will be made below on the point that the force for trapping a minute particle can be stably maintained strong by giving the plus spherical aberration to the converged beam. The present inventor has found that, by varying a condition of a light beam applied on a minute particle in a medium and calculating the trapping force in the direction of the optical axis in each case, the trapping force in the direction of the optical axis is strengthened when a plus spherical aberration is intentionally given to a converged beam applied on the minute particle in the medium. Then, as a result of intense examinations based on this founding, it is confirmed that, when a minute particle in a medium is irradiated with a converged beam having a plus spherical aberration, not only the strength of the trapping force in the direction of the optical axis is increased, but also the range over which the trapping force is exerted in the direction of the optical axis is expanded, and moreover, a sufficiently strong trapping force can be obtained even when the minute particle is present at a deep position inside the medium, that is, a position far from the converging optical system, and the distance the converged beam travels through the medium is considerably long.
Note that, in the minute particle optical manipulation apparatus according to the seventh aspect of the present invention, it is desirable that the axial chromatic aberration xcex94 of an observation light when using the trapping light in the converging optical system as a basis satisfied the following condition (1):
xe2x88x9210xe2x89xa6xcex94/Øxe2x89xa6xe2x88x920.12xe2x80x83xe2x80x83(1) 
where Ø is the size (for example, the diameter) of the minute particle.
Below the lower limit of the condition (1), the absolute value of the axial chromatic aberration xcex94 is too large so that the spherical aberration is generated in a large amount in a state in which the converging position of the converged beam is substantially coincident with the in-focus position and the trapping force of the minute particle becomes small undesirably. On the other hand, above the upper limit of the condition (1), the trapping force of the minute particle can not be stably maintained strong undesirably since the absolute value of the axial chromatic aberration xcex94 is too small so that the plus spherical aberration can not be given to the converged beam sufficiently. Note that, in order to exhibit the effects of the present invention more excellently it is more desirable that the lower limit of the condition (1) is set at xe2x88x925 and the upper limit at xe2x88x920.25.